Apparatus for the cultivation of microorganisms



March 27, 1962 R. R. FREEMAN APPARATUS FOR THE CULTIVATION OFMICROORGANISMS Original Filed Jan. 16, 1959 n w mm L K: Q /2 a A. r ah ag M 5 R m mu 0 1 W 8 k 7 m 0 m m .7 0 9 P w ATTORNEY United StatesPatent 3,027,305 APPARATUS FOR THE CULTIVATION OF MICROORGANISMS RobertR. Freeman, Pikesville, Md., assignor to the United The inventiondescribed herein may be manufactured and used by or for the Governmentof the United States of America for governmental purposes without thepayment to me of any royalty thereon.

This is a division of application Serial No. 787,312, filed January 16,1959, now Patent No. 2,970,088, granted January 31, 1961.

This invention relates to a novel fermentation apparatus and a novelfermentation process for the production of microorganisms.

Heretofore, various aerobic microorganisms were produced in a deep tankwhich contained a liquid nutrient, wherein air was supplied by means ofjets, spargers, draft tubes, etc., such that the mass transfer of oxygento the organism is dependent upon overcoming the resistances of thevarious film coeflicients. Usually it is necessary to provide anexpensive external power source in the form of mechanical agitators toovercome this resistance. Because of this aeration and agitation, thenutrients tend to foam since the nutrients are proteinaceous. Thisresults in the disadvantage of incurring considerable culture lossthrough the vent of the deep vat, or necessitating the addition ofchemical antifoams. Another disadvantage of the deep culture process isthe production of a very dilute suspension of microorganisms whichcauses the need for further concentration by evaporation, centrifuga ionor sedimentation.

In general, the object of my invention is to overcome thesedisadvantages by means of a simple and uncomplicated apparatus andprocess.

More specifically, the objects of my invention are as follows: (1) theuse of a solid nutrient medium in the process for the production ofmicroorganisms; (2) the use of a porous flexible material to provide alarge sur face area per unit volume for the support of the solidnutrient medium; and (3) a new apparatus designed to carry out theessential steps of the process which employs the use of a solid nutrienton a sponge or other type of material having a large surface area in theproduction microorganisms.

Other features and advantages of my invention will be come apparentduring the course of the following description with reference to theattached drawing.

Referring to the drawing which shows the entire layout wherein the airoperated motor 1 reciprocates piston rod 2 and piston 5 in a verticaldirection. The piston can be made of most any type of flexible materialbut is preferably made of a Teflon sheet. A steel back-up plate 4 isattached to the Teflon piston by means of nuts 6 to provide thenecessary rigidity to the plastic piston. The fermentor tank 3 containsa flexible porous support medium 7 which may be a natural or syntheticsponge. This sponge provides a very large surface area per unit volume.The sponge is supported by a perforated metal disc 9 which sits on ring10, which in turn is held by the bottom fermentor head 8. Entranceconduit 11a for fluids to the fermentor, and exit conduit 11b for fluidsfrom the fermentor, are supplied with manually operated valves 11c and11d, and electrically operated solenoid valves 12a and 12b. Thefermentor tank is closed at the top by its upper fermentor head 13,which contains a sealing ele- ICC ment 14 through which the piston rodor shaft slides, and a vacuum line 15 connected to a vacuum source forremoving any microorganisms which may get into the space between the topof the piston and the bottom of the upper fermentor head. Solenoidvalves (not shown) are contained in the air reversing switch 19 foroperating said switch. These solenoid valves along with the solenoidvalves 12a and 12b are required for properly timing the operation, andthey are activated by the pressure of the rotating cam 16 on themicroswitches 17. The cam is turned by a variable speed motor 18 andwhich is employed to select and to maintain the frequency ofv theoperating cycle.

Filters 20 and 23 are provided in order to prevent the microorganismsthat are being grown in the fermentor tank from escaping to theatmosphere and to prevent the contaminating organisms in the atmospherefrom entering the fermentor tank 3, the nutrient solution bottle 21a,the inoculum solution bottle 21b, and the wash solution bottle 210. Atemperature and humidity controlled air source 26 is connected to filter23 with manual valve 25 for introducing air to the fermentor tank.Bottles 21a, 21b and 210 containing nutrient, inoculum and washsolution, respectively, are connected to inlet conduit 11a by means of athree-way valve 24. A product receiver 22 is provided to catch and holdthe finished product.

In operation, the purpose of the piston is to aid in the feeding of thesolutions and air to sponge 7 and the removal therefrom of the excesssolutions and spent air by its reciprocating action on the sponge. Thespeeds of each of these operations are controlled and can be varied bythe operation of the variable speed motor which in turn controls thecam, the limit switches and the solenoid valves.

My novel process will now be explained in connection with theaforementioned apparatus described above, with out, however, limiting myprocess in the employment of said apparatus.

The process is started with the sponge being compressed by the piston.The variable speed motor is started and its speed is regulated so thatit will tend to suck nutrient at the proper temperature from thenutrient bottle into the fermentor tank and sponge. This flow can beaided if the nutrient bottle is situated higher in level than thefermentor tank so that the liquid can flow by gravity. Manual valve isopened but manual valve 11d is closed so that the operation of thesolenoid valves in this operation is of no consequence. After the spongeis saturated with nutrient, manual valve 110 is closed and manual valve11d is opened so that the piston on the downward stroke can remove theexcess nutrient. The piston is raised, the variable speed motor isstopped and valve 11d is closed. The nutrient is allowed to cool at thispoint in order to solidify the nutrient. This cooling can be increasedby the use of a cooling jacket, .if desired.

After the nutrient has been solidified as a coating on the poroussurfaces throughout the sponge, the variable speed motor is started,thereby activating the piston which compresses the sponge. Valve 110 isthen opened and the three-way valve 24 is adjusted so that inoculum isdrawn into the fermentor tank on the upward swing of the piston. Whenthe sponge is saturated the excess is removed using the same procedureas used in eliminating the excess nutrient. At this point both manualvalves 11c and 11d are opened, the three-way valve is closed and thevalve 25 is opened, which allows air at the proper temperature to enterthe fermentor tank on the upswing of the piston. After the tank isfilled with air, the solenoid valve 12a closes and the solenoid valve12b opens on the downward swing of the piston in order to force the airfrom the tank. This "breathing period is maintained until the desiredamount of microorganisms are produced. At this point the air supply isclosed and wash solution is fedto the tank in the same manner asdescribed for the nutrient and inoculum. When the sponge has beensaturated, the speed of variable speed motor is increased in order toproduce a violent agitation, thereby forcing the microorganisms and washsolution from the tank into the receiver where it is collected forfurther purification steps if desired.

The process as described above is extremely valuable in the productionof various types of aerobic microorganisms in the laboratory. Theprocess of course can be used for production of anaerobic microorganismsby eliminating the addition of air to the system, and by controlling thetemperature by other well known means, for example, the use of a heatingjacket. It is evident that the process with a few modifications such asproviding a time controller to operate the variable speed motor, andreplacing the manual valves with solenoid valves, can be converted intoan uninterrupted cyclic process for the production of any one type ofmicroorganism.

It is also possible that a liquid nutrient can be employed in lieu of asolid nutrient; however, the use of a liquid nutrient would not be asadvantageous as explained earlier.

I claim:

l. A fermentor tank having a movable piston and sponge therein, a shaft,power means, said shaft being connected to said power means and to saidpiston in order to move said piston; said tank comprising a cylindricalsidewall, upper fermentor head attached to said sidewalls for closingoff the top of the tank, and having an aperture in the center for saidshaft to extend therethrough, a sealing element attached to said upperfermentor head through which a portion of said shaft slides, saidsealing element having a vacuum source connected thereto to remove bysuction any microorganisms which may get into the space between the topof the piston and the bottom of said upper fermentor head, a fermentorbottom head removably attached to said cylindrical side wall and havingtwo apertures therethrough for connecting with two conduits, a ringsupport held by said bottom head, and a perforated plate held by saidring which is used to support said sponge.

2. A fermentor system for the production of microorganisms whichcomprises a fermentor tank, a solid support medium which provides alarge surface area per unit volume contained in said tank, conduit lineswith manual and electrical solenoid valves for supplying and removingmaterials to and from said tank, power means connected to said tank forforcing the reactants to the tank and the products from said tank, andcontrol means connected to said power means andto said solenoid valvesfor producing the desired feed and removal rate of the reactants andproducts to and-from said fermentor tank; said power means comprising afluid pressure means, and a piston means which is positioned within saidfermentor tank.

3. A fermentor system as set forth in claim 2 wherein said solid supportmedium comprises a porous, flexible, spongy type of material which isused to support the nutrient and inoculum substances.

4. A fermentor system as set forth in claim 2 in which said controlmeans comprises a motor driven cam with a series of limit switches whichactivate said power means and said solenoid valves.

5. A fermentor system as set forth in claim 2 in which said piston meanscomprises a piston made of a plastic sheet, a steel back-up plateconnected to said plastic piston to provide rigidity, and a shaftconnecting the back-up plate and plastic piston with said fluid pressuremeans.

6. A fermentor system as set forth in claim 2 wherein said fluidpressure means comprises an air operated motor and an air reversingswitch.

,7. A fermentor system as set forth in claim 6 wherein said solidsupport medium comprises a porous, flexible, spongy type of materialwhich is used to support the nutrient and inoculum substances.

References Cited in the file of this patent UNITED STATES PATENTS454,586 Bachmann s June 23, 1891

1. A FERMENTOR TANK HAVING A MOVABLE PISTON AND SPONGE THEREIN, A SHAFT,POWER MEANS, SAID SHAFT BEING CONNECTED TO SAID POWER MEANS AND TO SAIDPISTON IN ORDER TO MOVE SAID PISTON; SAID TANK COMPRISING A CYLINDRICALSIDEWALL, UPPER FERMENTOR HEAD ATTACHED TO SAID SIDEWALLS FOR CLOSINGOFF THE TOP OF THE TANK, AND HAVING AN APERTURE IN THE CENTER FOR SAIDSHAFT TO EXTEND THERETHROUGH, A SEALING ELEMENT ATTACHED TO SAID UPPERFERMENTOR HEAD THROUGH WHICH A PORTION OF SAID SHAFT SLIDES, SAIDSEALING ELEMENT HAVING A VACUUM SOURCE CONNECTED THERETO TO REMOVE BYSUCTION ANY MICROORGANISMS WHICH MAY GET INTO THE SPACE BETWEEN THE TOPOF THE PISTON AND THE BOTTOM OF SAID UPPER FERMENTOR HEAD, A FERMENTORBOTTOM HEAD REMOVABLY ATTACHED TO SAID CYLINDRICAL SIDE WALL AND HAVINGTWO APERTURES THERETHROUGH FOR CONNECTING WITH TWO CONDUITS, A RINGSUPPORT HELD BY SAID BOTTOM HEAD, AND A PERFORATED PLATE HELD BY SAIDRING WHICH IS USED TO SUPPORT SAID SPONGE.